Human Total EGFR DuoSet IC ELISA
R&D Systems, part of Bio-Techne | Catalog # DYC1854-2
Key Product Details
Assay Type
Solid Phase Sandwich ELISA
Assay Range
31.2-2000 pg/mL
Sample Type
Cell Lysates
Reactivity
Human
Human Total EGFR DuoSet IC ELISA Features
- Optimized capture and detection antibody pairings with recommended concentrations save lengthy development time
- Development protocols are provided to guide further assay optimization
- Assay can be customized to your specific needs
- Available in 2, 5, and 15- (96-well) plate pack sizes
- Economical alternative to Western blot
Product Summary for Human Total EGFR DuoSet IC ELISA
This DuoSet IC ELISA contains the basic components required for the development of sandwich ELISAs to measure total EGFR in cell lysates. An immobilized capture antibody specific for EGFR binds both phosphorylated and unphosphorylated EGFR. After washing away unbound material, a biotinylated detection antibody is used to detect both phosphorylated and unphosphorylated protein, utilizing a standard Streptavidin-HRP format.
Product Specifications
Assay Format
96-well strip plate (sold separately)
Sample Volume Required
Cell lysates (100 µL)
Detection Method
Colorimetric ELISA - 450nm (TMB)
Conjugate
Biotin
Specificity
Label
HRP
Scientific Data Images for Human Total EGFR DuoSet IC ELISA
Standard Curve
Figure 1. The Human Total-EGF R DuoSet IC ELISA is more sensitive than immunoprecipitation (IP)-Western blot analysis
Lysates prepared from the human epidermoid carcinoma cell line, A431, were diluted in series and analyzed by (A) IP-Western blot and (B) R&D Systems’ Human Total-EGF R DuoSet IC ELISA (Catalog # DYC1854). IPs were performed using anti-EGF R polyclonal antibody (Catalog # AF231) Immunoblots were incubated with a biotinylated anti-EGF R polyclonal antibody (Catalog # BAF231) to detect total EGF R. Blots were incubated with Streptavidin-HRP (Catalog # DY998) followed by chemiluminescent detection. Human EGF R can be detected by the Human Total-EGF R DuoSet IC ELISA by using approximately 200 to 500 times less lysate than is needed for a conventional IP-Western blot.
Figure 2. The Human Total-EGF R DuoSet IC ELISA measures the relative level of EGF R
Lysates were prepared from A431 cells, human immortalized fibroblast cells (CCD1070SK), and human breast adenocarcinoma cells (SkBr3). ELISA analysis was done using 125 ng (A431) and 2000 ng (CCD1070SK and SkBr3) of lysate. IP-Western blot analysis (inset) was done using 5 μg (A431) and 100 μg (CCD1070SK and SkBr3) of lysate. The IP-Western blot was performed as described in Figure 1. The smaller band visible in IP-Western blot is an immature EGF R glycosylation intermediate.Kit Contents for Human Total EGFR DuoSet IC ELISA
- Capture Antibody
- Conjugated Detection Antibody
- Calibrated Immunoassay Standard or Control
- Streptavidin-HRP
Other Reagents Required
PBS: (Catalog # DY006), or 137 mM NaCl, 2.7 mM KCl, 8.1 mM Na2HPO4, 1.5 mM KH2O4, pH 7.2 - 7.4, 0.2 µm filtered
Wash Buffer: (Catalog # WA126), or equivalent
Lysis Buffer*
IC Diluent*
Blocking Buffer*
Substrate Solution: 1:1 mixture of Color Reagent A (H2O2) and Color Reagent B (Tetramethylbenzidine) (Catalog # DY999)
Stop Solution: 2 N H2SO4 (Catalog # DY994)
Microplates: From Costar EIA Plate (Costar Catalog # 2592) or R&D Systems (Catalog # DY990), or equivalent
Plate Sealers: ELISA Plate Sealers (Catalog # DY992), or equivalent
*For the Lysis Buffer, IC Diluent, and Blocking BUffer recommended for a specific DuoSet ELISA Development Kit, please see the product
Preparation and Storage
Shipping
The product is shipped at ambient temperature. Upon receipt, store it immediately at the temperature recommended below.
Stability & Storage
Store the unopened product at 2 - 8 °C. Do not use past expiration date.
Background: EGFR
EGFR (epidermal growth factor receptor), also known as HER-1, ErbB1, or ErbB, is a transmembrane receptor tyrosine kinase that binds a subset of the EGF family ligands including EGF, Amphiregulin, TGF-alpha, Betacellulin, Epiregulin, HB-EGF, and Epigen. Ligand binding induces EGFR homodimerization as well as heterdimerization with ErbB2, resulting in kinase activation, tyrosine phosphorylation and cell signaling. EGFR can also be recruited to form heterodimers with the ligand-activated ErbB3 or ErbB4. EGFR signaling regulates multiple biological functions including cell proliferation, differentiation, motility, and apoptosis. EGFR is overexpressed in a wide variety of tumors and is the target of several anti-cancer drugs. Soluble receptors consisting of the extracellular ligand binding domain are generated by alternate splicing in human and mouse.
Long Name
Epidermal Growth Factor Receptor
Alternate Names
EGF R, ErbB, ErbB1, HER-1
Gene Symbol
EGFR
Additional EGFR Products
Product Documents for Human Total EGFR DuoSet IC ELISA
Product Specific Notices for Human Total EGFR DuoSet IC ELISA
For research use only
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